Redox biochemistry is fundamental to life. The energy needs of complex organisms require vast amounts of ATP. The supply of ATP depends heavily on redox chemistry, as it is driven by changes in free energy associated with electron or hydrogen transfers (Frein et al., (2005) Biochem Pharmacol 70:811-823). Technically, redox, shorthand for reduction/oxidation, describes all chemical reactions in which the oxidation state of atoms changes. In simpler terms, oxidation describes the loss of electrons by a molecule, atom, or ion and reduction describes the gain of electrons by the same.
Redox signaling is the concept that electron-transfer processes play a key messenger role in biological systems. At the heart of redox signaling are the so-called reactive oxygen species (ROS), including oxygen radicals (e.g., O2.− and OH.) and also nonradical derivatives of O2 (H2O2). The discovery of reactive nitrogen species expanded this term to “reactive oxygen and nitrogen species” (RONS). Free radicals contain one or more unpaired electrons. Since molecules seek to be balanced, that is to have an equal number of protons and electrons, the unpaired electron spins of these radicals make them highly reactive.
RONS are produced continuously by the mitochondria (O2.−, OH. and H2O2,) of most cells and also by cytochrome P450 (O2.−, and H2O2), macrophages, (O2−, H2O2, and NO) and peroxisomes (H2O2) (Klaunig, et al., (2004) Annu Rev Pharmacol Toxicol 44:239-267; Genestra, M., (2007) Cell Signal 19:1807-1819). During mitochondrial oxidative metabolism, about 5% of oxygen is converted primarily into O2, whereas 95% of it is reduced to water. Given the high reactivity of RONS, it is not surprising that the cell has invested heavily into an antioxidant defense system to contain RONS. This defense system includes: (a) classic antioxidant enzymes, such as superoxide dismutase (SOD), catalase, glutathione (GSH) peroxidase, glutaredoxin, and thioredoxin, which are distributed in mitochondria, peroxisomes, and cytoplasm; (b) Nonclassic antioxidant enzymes, for example, heme oxygenase-1; (c) Phase II detoxifying enzymes, recently shown to be protective, such as GSH reductase, NQO1, and GSH transferasel; and (d) nonenzymatic antioxidants, such as vitamins E and C, GSH, and catechins.
For many years, it has been widely assumed that all RONS are bad for the cell. Consequently, research efforts have been generally focused on suppressing RONS, hoping to prevent or even reverse RONS-related biological damage. Reactive oxygen and nitrogen species, however, have what can be termed ‘multiple biological personalities:’ at low concentrations they protect the cell; at higher concentrations they can damage many biological molecules, such as DNA, proteins, and lipids; and yet, they can also help prevent cancer by initiating the death of the transformed cell.
Recently an important conceptual distinction has become clear with regard to the roles reactive oxygen and nitrogen species play in cellular physiology. (Frein et al., (2005) Biochem Pharmacol 70:811-823); Halliwell, B., (2007) Biochem J 401:1-11). First, it has been well-established that there exists a network of redox-based regulatory mechanisms that are often quite relevant to carcinogenesis, but which are not necessarily pathophysiological. Second, it is also clear that disturbed redox equilibrium is indeed pathophysiological and such disturbances have been described for years as ‘oxidative stress.’ These findings have led to the delineation between redox signaling and oxidative stress. Redox signaling embraces a reversible phase of physiological regulatory reactions occurring over short time periods where the signal is passed via the addition and loss of electrons. These regulatory reactions relate primarily the main cellular redox systems, e.g., GSH, ascorbate, vitamin E, lipoic acid, NADPH, or NADH. In this type of signaling, the oxidative reactions, which often lead to posttranslational protein modification, are returned to the resting state by reductive pathways. Such posttranslational modifications include glutathiolation, S-nitrosylation, methionine sulphoxidation, and oxidations with disulfide formation. In contrast, oxidative stress denotes a persistent and often irreversible oxidative shift that characterizes a pathophysiological state. Oxidative stress has been defined as an imbalance between oxidants and antioxidants in favor of the former, resulting in increased cellular levels of RONS. Oxidative stress is implicated in the pathogenesis of several diseases including cancer, inflammatory disorders, cardiovascular and neurodegenerative disorders, sepsis, reperfusion damage, rheumatoid arthritis, osteoarthritis, and diabetes.
There is significant evidence for a role of RONS in cancer. Specific activities where RONS have been implicated include genotoxicity, promotion of transformed cell growth and angiogenesis, as well as the regulation of apoptosis. For example, persistent oxidative stress has been suggested in Toyokuni et al., (1995) to contribute to oncogene activation, genomic instability, chemotherapy resistance, and even invasion and metastasis. (Toyokuni et al., (1995) FEBS Lett 358: 1-3). Nuclear factor-kB (NF-kB), MAPK cascades as well as GSH and related antioxidant pathways are suggested to be the mediators of such RONS-related activity. Chronic inflammation, which is widely considered to be connected to carcinogenesis, is another source of RONS. The linkage of RONS generated by inflammation to cancer has also been postulated. Similarly, the hypoxiainducible factor-1a (HIF-1a) is linked to cancer through its regulation by RONS. (Pouyssegur et al., (2006) Biol Chem 387:1337-1346). In particular, RONS signaling can account for the high levels of HIF-1a in normoxic areas of tumors. Hypoxia-inducible factor promotes survival in low oxygen conditions, like those encountered in many cancerous tumors, by upregulating an array of hypoxia-induced genes, including the vascular endothelial factor, which promotes angiogenesis. Finally, RONS have been associated with the induction of apoptotic and necrotic cell death, the specific outcome depending on, inter alia, the cellular levels of RONS.
As noted in Rigas, B., and Sun, Y., (2008), altering the redox status of a cancer cell can result in the death of that cell. (Rigas, B., and Sun, Y., (2008) British J. of Cancer, 98:1157-1160, the contents of which are expressly incorporated by reference herein). In particular, Rigas and Sun were able to establish that increased production of RONS leads to oxidative stress and apoptosis in cancer cells. Thus, intervention in the redox state of a particular cancer cell provides a strategy for treatment and/or prevention of that cancer. In fact, various anticancer agents are already well-known to induce the production of RONS and induce cell death through oxidative stress, including the topoisomerase inhibitor etoposide (Oh et al., (2007) Mol Cancer Ther 6:2178-2187), arsenic trioxide (Nakagawa et al., (2002) Life Sci 70:2253-2269), and cisplatin (Berndtsson et al., (2007) Int J Cancer 120:175-180).
For a long time, reactive oxygen species have been considered harmful mediators of inflammation owing to their highly reactive nature. However, emerging findings suggest that ROS can be anti-inflammatory and prevent autoimmune responses, thus challenging existing dogma. For instance, ROS produced by the phagocyte NADPH oxidase (NOX2) complex might be produced as a mechanism to fine-tune the inflammatory response. (Hultqvist et al., (2009) Trends Immunol. 30(5):201-208). To illustrate this point further, recent evidence suggests that NO and its redox derivatives may protect joints affected by osteoarthritis, a degenerative disease involving chondrocytes and cartilage (Abramson, (2008) Arthritis Res Ther. 10 Suppl 2:52). NO and its derivatives have a similarly protective involvement in nociception and pain, which may contribute to the functional disability of osteoarthritis. A similar understanding has been developed for other inflammation-related clinical entities. The critical role of inappropriate inflammation is becoming accepted in many diseases, including cardiovascular diseases, inflammatory and autoimmune disorders, neurodegenerative conditions, infection and cancer (Smith, G. and Missailidis, S., (2004) Journal of Inflammation 1:3 and Zhang, Z. and Rigas, B., (2006) Int J Oncol. 29(1):185-92).
There is a continuing need in the art to identify new therapies for RONS-related pathologies, e.g., cancer, inflammatory disorders, cardiovascular disorders, neurodegenerative disorders, sepsis, reperfusion damage, rheumatoid arthritis, osteoarthritis, and diabetes, including a need for new agents that function via the inducing the production of RONS. The present invention provides an entirely new class of such RONS-generating agents.